Figure 7. NR4A2 was regulated by NF-κB directly in H₂O₂-induced autophagy and apoptosis of CSCs.

CSCs were challenged with 500 μM H₂O₂ for 5 h. (A) Representative immunofluorescent images showed the translocation of P65 in CSCs. (B) Change of phosphorylated P65 were detected by western blot. (C) The caspase3 activity was detected by Caspase3 Colorimetric Assay kit. (D) CSCs were pretreated with NF-κB inhibitor bay11-7082 (10 μM) for 30 min, then treated with H₂O₂ for 5 h, western blot showed the changes of phosphorylated IκBα, cleaved PARP1, LC3-II and P62 in CSCs. (E–G) The changes of NR4A2 in CSCs treated with bay11-7082 combined with H₂O₂ were detected by immunofluorescent assay (E), the qPCR (F), western blot (G) and respectively. (H) Luciferase reporter plasmid containing the NR4A2 promoter was co-transfected with Flag-P65-AMP overexpression plasmid in HEK293 cells for 24 h followed by dual luciferase activity assay. ctr, control. *P < 0.05; **P < 0.01; n=3.